Analysis of Caffeine in Coffee
Kate Penrod, Nikki DeLuca, Christina
Robinson, and Jess Bases
Purpose
• Quantify caffeine in
brewed coffee
– Varying strengths
• Extract caffeine from
grounds
• Maxwell House:
Regular and Decaf.
Background
•
•
•
•
Caffeine in food/drink
CNS stimulant
Schedule V drug
High levels
–
–
–
–
Restlessness
Irritability
Headaches
Heart complications
Methods
• HPLC
– Liquid
• Separation
– Liquid
• IR
– Solid and liquid
• SFE
– Solid
• UV/Vis
– Disabled
HPLC
• Hypothesis
• Samples
–
–
–
–
10-50ppm caffeine standards
Strong caffeinated/decaffeinated
Regular caffeinated/decaffeinated
Weak caffeinated/decaffeinated
• Dilution of coffee samples
Sample
Conc [ppm] Retention Time
Area
Standard
10
2.346
1026491
Standard
20
2.323
1360973
Standard
30
2.314
1612710
Standard
40
2.305
1915989
Standard
50
2.302
2330806
Weak DC 1:2
46.55369153
2.266
1436697
Reg DC 1:5
77.88165933
2.293
1193080
Str DC 1:5
227.381635
2.283
2139010
Weak C 1:5
180.6469498
2.294
1843306
Reg C 1:10
321.8074968
2.313
1718385
Str C 1:25
548.2456002
2.310
1394082
Organic Separation
• Sample Preparation (Dr. Peterman’s Coffeemaker)
– Regular Caffeinated/ Decaffeinated
• 1 Tbsp. grounds, ¾ cup water
– Strong Caffeinated/ Decaffeinated
• 3/2 Tbsp. grounds, ¾ cup water
– Weak Caffeinated/ Decaffeinated
• ½ Tbsp. grounds, ¾ cup water
Organic Separation
• First Method
– Adapted from Dr. Halligan’s Organic I Lab Manual
– Extraction of Caffeine using
Dichloromethane
– Large emulsion layer,
not a clear separation
Organic Separation
• Revised Method
– Added Na2CO3 which increased
caffeine’s solubility in aqueous layer
– Separation layer more pronounced
– Dried with Anhydrous MgSO4
– Vacuum under reduced pressure
Mass Quantification
Appearance
Mass of Product
Regular Caffeinated
Yellow/brown liquid
0.2753g
Weak Caffeinated
Light brown liquid
0.2731g
Strong Caffeinated
Yellow/ brown solid,
black spots
0.2892 g
Regular Decaffeinated
Yellow/brown liquid
0.2006 g
Weak Decaffeinated
Light brown liquid
0.2001g
Strong Decaffeinated
Yellow/ brown liquid
0.2296 g
Extracted Caffeine IR Characterization
Substituted Alkene
Extracted Caffeine IR Characterization
O – H stretch
C=O
C=N
C=O
CH3
C-N
C=N
Substituted Alkene
Substituted Alkene
Regular Decaff
Strong Decaff
C=O
O – H stretch
C=N
Substituted Alkene
Weak Decaff
Extracted Caffeine IR Characterization
O – H stretch
O – H stretch
C=O
CH3
C=N
CH3
C=O
C-N
C=N
C-N
Substituted Alkene
Substituted Alkene
Regular Caffeinated
Strong Caffeinated
C=N
O – H stretch
CH3
C-N
Substituted Alkene
Weak Caffeinated
The Devil’s Instrument
a.k.a. SFE
The Devil’s Instrument
a.k.a. SFE
• 4 Samples
– “Pure” coffee grounds
– Post brewing coffee grounds
•
•
•
•
1 gram of each sample
10 minute runs
End mass indications
IR results
SFE Data
Sample
Percent Yield
Pure Caffeinated
5.186 %
Pure Decaf.
6.286 %
Brewed Caffeinated
1.185 %
Brewed Decaf.
3.576 %
SFE Extraction IR Analysis
UV/Vis
• Attempted to use the DRA-600
– Solid Analysis
• Missing part!
• Call EVERYONE
– Manufacturer
– Distributor
– Delivery Guy
• Piece was backordered until May 20th
• Put on hold INDEFINITELY.
Sources of Error
• Organic Method
– Impure caffeine for mass quanitification
– Transfer between different glassware
– Incomplete drying in extraction (O-H stretch peak)
• HPLC
– Day 1: Low pressure, purge lines
– Day 2: Invalid date/time error, inconsistent
– Day 3: Consistent results, ran all samples
Sources of Error (Part 2)
• SFE
– Leaks in the line
– Issues with the amount of sample
– Needed constant supervision
• UV/Vis
– Missing part
– Phone tag
– Wasted 2 whole lab periods
Sources of Error (Part 3)
Jerry Sainsbury – Sales Representative
Thermo Fisher Scientific
Conclusions
• HPLC:
• Strong decaf. was comparable to weak caf.
• Weak and regular brews – 4:1 caf. vs. decaf.
• Strong brew – 5:2 caf. vs. decaf.
• Organic Method:
• Qualitative data
• Erroneous quantitative data
• Masses very similar
• SFE:
• Low percent yields
• Higher yield from decaf
• Did not extract pure caffeine
• May be caffeine plus side products
• If UV/Vis hadn’t been in the initial method, further research would have been
conducted
Further Research
• Organic Method
– Develop a new or revised method to extract pure
caffeine in its solid form as was done in Dr.
Halligan’s Organic I Lab
• IR
– Analyze liquid only
– Create calibration curve
• HPLC
– Compare different coffee makers and brands
Further Research (Part 2)
• SFE
– Brew remaining grounds, analyze with HPLC
– Determine other components of extracted
grounds
• UV/Vis
– Obtain calibration disc and sample holder
– Develop SOP for DRA-600